Collagen cross-linking is required for synthesis of functional collagen fibrils in vivo. After synthesis of collagen monomers, the initial reaction in the biosynthesis of collagen cross-links is formation of aldehyde cross-link intermediates and is catalyzed by lysyl oxidase. This report summarizes research relating to embryonic chick bone lysyl oxidase and collagen cross-linking. The enzyme has been purified to homogeneity on SDS-acrylamide electrophoresis and has a molecular weight of approximately 63,000 daltons. Principal findings were: (1) The highly purified enzyme had high activity with reconstituted, native collagen fibrils and low activity with either collagen monomers or "non-striated" fibrils. (2) Enzyme dependent synthesis of the two Schiff base cross-links, hydroxylysinonorleucine (HLNL) and hydroxylysino-hydroxynorleucine (DHLNL), occurred in the fibrils. (3) With time the concentration of HLNL rose and reached a plateau. DHLNL rose more rapidly and decreased to low levels. (4) A new uncharacterized compound with molecular weight higher than DHLNL appeared as the concentration of DHLNL decreased. (5) The highly purified enzyme catalyzed formation of aldehyde intermediates in elastin and in both lysyl and hydroxylysyl residues in collagen. (6) Synthesis of histidinohydroxymerodesmosine was demonstrated with highly purified enzyme in vitro. The purpose of the present proposal is to continue to study lysyl oxidase and the biosynthesis of collagen cross-links with highly purified lysyl oxidase and collagen in vitro. The principal aims are to: (1) Define the collagen substrate more precisely by determining whether the protease-sensitive, carboxyterminal non-helical region is present. (2) Determine the location of the enzyme binding site to the helical portion of collagen molecule. (3) Loocate the sites of enzyme cross-linking in collagen by studying cross-links in the collagen cyanogen bromide peptides and by using model collagens such as (alpha 1)3 and (alpha 2)3 as substrates and then characterizing the reaction products. (4) Determine whether type II (cartilage) collagen is a substrate for the same enzyme as type I collagen with similar kinetics. (5) Characterize the new high molecular weight compound derived from DHLNL during cross-linkin (Text Truncated - Exceeds Capacity)